Testis specific 10 (TSGA10) was originally identified as a testis-specific protein and tumor-associated antigen in a number of cancer types. In this study, we found that down-regulation of TSGA10 was associated with increased malignancy and clinical features of esophageal squamous cell carcinomas (ESCCs). Moreover, increased expression of TSGA10 inhibited, while its knockdown promoted, tumor formation in vivo in nude mice. At the 3′UTR of the TSGA10 gene we identified two binding sites for microRNA-577 (miR-577). Further investigation demonstrated that expression levels of miR-577 and TSGA10 were negatively correlated to each other in ESCC cell lines and tumor samples. Moreover, manipulation of miR-577 and TSGA10 expression confirmed that miR-577 can regulate TSGA10 and in turn affect cell proliferation in vitro. Additionally, with flow cytometry and manipulation of the mir-577/TSGA10 axis, it was found that mir-577/TSGA10 axis influenced the growth of ESCC through regulating the G1-S phase transition. We also obtained evidence to establish that mir-577/TSGA10 axis activation was always accompanied by inactivation of the p53 pathway or the Rb pathway or both, thus, the latter two pathways are obligatory for progression of ESCCs with mir-577/TSGA10 axis activation. In addition, we found that such an interactive pathway in regulating cancer cell proliferation was restricted to a few cancer types including ESCC, but not uniformly applicable to other cancer types. This newly discovered regulatory mechanism provides a new dimension for ESCC diagnosis and therapy.